title: Development of fluorescent substrates for microsomal epoxide hydrolase and application to inhibition studies.
abstract: The microsomal epoxide hydrolase (mEH) plays a significant role in the metabolism of numerous xenobiotics. In addition, it has a potential role in sexual development and bile acid transport, and it is associated with a number of diseases such as emphysema, spontaneous abortion, eclampsia, and several forms of cancer. Toward developing chemical tools to study the biological role of mEH, we designed and synthesized a series of absorbent and fluorescent substrates. The highest activity for both rat and human mEH was obtained with the fluorescent substrate cyano(6-methoxy-naphthalen-2-yl)methyl glycidyl carbonate (11). An in vitro inhibition assay using this substrate ranked a series of known inhibitors similarly to the assay that used radioactive cis-stilbene oxide but with a greater discrimination between inhibitors. These results demonstrate that the new fluorescence-based assay is a useful tool for the discovery of structure-activity relationships among mEH inhibitors. Furthermore, this substrate could also be used for the screening chemical library with high accuracy and with a Z' value of approximately 0.7. This new assay permits a significant decrease in labor and cost and also offers the advantage of a continuous readout. However, it should not be used with crude enzyme preparations due to interfering reactions.
authors.affiliation: Department of Entomology and Cancer Center, University of California - Davis, 95616, USA. chmorisseau@ucdavis.edu
authors.forename: Christophe
authors.initials: C
authors.lastname: Morisseau
authors.forename: Maud
authors.initials: M
authors.lastname: Bernay
authors.forename: Aurélie
authors.initials: A
authors.lastname: Escaich
authors.forename: James R
authors.initials: JR
authors.lastname: Sanborn
authors.forename: Jozsef
authors.initials: J
authors.lastname: Lango
authors.forename: Bruce D
authors.initials: BD
authors.lastname: Hammock
mesh_terms: D000818:Animals; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D004851:Epoxide Hydrolases / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D051381:Rats; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D004851:Epoxide Hydrolases